Regurgitation of locusts dedicates chemical defense against predators

Regurgitation in insects is generally regarded as having digestive and defensive functions. Despite a significant aversion of locust regurgitation to some predators, the defensive mechanisms against predators remain unclear. Herein, we employ the locust-ant as a prey-predator system to investigate the defensive mechanism of locust regurgitation by behavioral bioassays, chemical analyses, and electrophysiological experiments. We found that locusts can use their regurgitation in response to a wide range of predators, including fire ants, mantises, and sparrows. Locust regurgitation is mainly secreted from the crop and salivary glands (SG). The colony and individual foraging behavior of ants demonstrated that locust regurgitation and mealworms smeared with the regurgitation significantly repelled the ants' attack. A combination of chemical analysis and behavior assays confirmed that small-molecule compounds, rather than proteins in locust regurgitation, exerted the defensive effect. Among the 907 compounds in the regurgitation, only the mixture of diethyl glutarate (DG) from the SG and 3-methylindole (3MI) and 2-(formylamino)benzoic acid (FBA) from the crop can deter predation of the ants. Furthermore, the mixture of three compounds mediates defense and aversion by stimulating ant gustation based on electrophysiological trials. Our findings reveal molecular evidence of locust regurgitation for chemical defense, offering a pivotal compound combination mediating the interactions in prey-predator systems.